Protecting osmolytes are responsible for the kidney medulla's extraordinary ability to cope with intracellular urea concentration as high as 1.5M. These small organic molecules are common in cells of many tissues, and an important part of their action in kidney is to stabilize intracellular proteins against the deleterious effects of urea. Besides being essential for our survival, imbalances in osmolyte levels play key roles in such conditions as polycystic kidney disease, diabetes mellitus, and brain edema. Though many biological roles of osmolytes arise from their solvation of proteins, it is unknown how protein solvation facilitates these roles. This gap in knowledge prevents a complete understanding of osmolyte effects and their roles in normal and disease states. Our long-term goal is to understand how interactions among osmolytes, water, and biomolecules give rise to osmotic stress response on the one hand, and disease on the other. Using measurements of the transfer free energy of protein side-chain and peptide backbone groups (GTFEs) from water to osmolyte solution, we recently made the remarkable discovery of how to predict the energetics of protein-stability in osmolytes. Our aims are to consolidate and use this ability to determine the underlying forces responsible for protein stability, and to the predict energetic effects of osmolytes on contraction and accretion of structure in denatured ensembles. We will extend our use of GTFEs to enable predictions of protein-protein interaction free energies, to better understand how fluctuating osmolyte concentrations affect key protein-protein interactions vital to cellular responses, and determine the extent to which kidney osmolytes act synergistically, negatively, or independently in affecting the properties of proteins. We aim to merge our ability to predict the energetics of protein stability and solvation effects with Kirkwood-Buff approaches that structurally relate water*osmolyte*protein interaction with the energetics, to give a considerably more detailed mechanistic understanding of protein solvation than currently exists. Relevance: This project will lead to a better understanding of how osmolytes protect proteins from unfolding under harsh condition, and how imbalances in osmolyte levels can contribute to the pathology of such conditions as polycystic kidney disease, diabetes mellitus, and brain swelling. Our work will have practical applications in the pharmaceutical industry for stabilization of vaccines and protein/ peptide drugs.